| Autor: |
Manosas, Maria, Camunas-Soler, Joan, Croquette, Vincent, Ritort, Felix |
| Předmět: |
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| Zdroj: |
Nature Communications; 8/21/2017, Vol. 8 Issue 1, p1-12, 12p |
| Abstrakt: |
Most DNA processes are governed by molecular interactions that take place in a sequence-specific manner. Determining the sequence selectivity of DNA ligands is still a challenge, particularly for small drugs where labeling or sequencing methods do not perform well. Here, we present a fast and accurate method based on parallelized single molecule magnetic tweezers to detect the sequence selectivity and characterize the thermodynamics and kinetics of binding in a single assay. Mechanical manipulation of DNA hairpins with an engineered sequence is used to detect ligand binding as blocking events during DNA unzipping, allowing determination of ligand selectivity both for small drugs and large proteins with nearly base-pair resolution in an unbiased fashion. The assay allows investigation of subtle details such as the effect of flanking sequences or binding cooperativity. Unzipping assays on hairpin substrates with an optimized flat free energy landscape containing all binding motifs allows determination of the ligand mechanical footprint, recognition site, and binding orientation. [ABSTRACT FROM AUTHOR] |
| Databáze: |
Complementary Index |
| Externí odkaz: |
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